Triple-modal imaging of stem-cells labeled with multimodal nanoparticles, applied in a stroke model

Helio Rodrigues da Silva; Javier Bustamante Mamani; Mariana Penteado Nucci; Leopoldo Penteado Nucci; Andrea Tiemi Kondo; Daianne Maciely Carvalho Fantacini; Lucas Eduardo Botelho de Souza; Virginia Picanço-Castro; Dimas Tadeu Covas; José Mauro Kutner; Fernando Anselmo de Oliveira; Nelson Hamerschlak; Lionel Fernel Gamarra

doi:10.4252/wjsc.v11.i2.100

Triple-modal imaging of stem-cells labeled with multimodal nanoparticles, applied in a stroke model

World J Stem Cells. 2019 Feb 26;11(2):100-123. doi: 10.4252/wjsc.v11.i2.100.

Authors

Affiliations

¹ Hospital Israelita Albert Einstein, São Paulo 05651-900, Brazil.
² LIM44, Hospital das Clinicas HCFMUSP, Faculdade de Medicina, Universidade de São Paulo, São Paulo 05403-010, Brazil.
³ Centro Universitário do Planalto Central, Brasilia 72445-020, Brazil.
⁴ Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, São Paulo 05403-010, Brazil.
⁵ Hospital Israelita Albert Einstein, São Paulo 05651-900, Brazil. lgamarra@einstein.br.

Abstract

Background: Mesenchymal stem cells (MSCs) have been widely tested for their therapeutic efficacy in the ischemic brain and have been shown to provide several benefits. A major obstacle to the clinical translation of these therapies has been the inability to noninvasively monitor the best route, cell doses, and collateral effects while ensuring the survival and effective biological functioning of the transplanted stem cells. Technological advances in multimodal imaging have allowed in vivo monitoring of the biodistribution and viability of transplanted stem cells due to a combination of imaging technologies associated with multimodal nanoparticles (MNPs) using new labels and covers to achieve low toxicity and longtime residence in cells.

Aim: To evaluate the sensitivity of triple-modal imaging of stem cells labeled with MNPs and applied in a stroke model.

Methods: After the isolation and immunophenotypic characterization of human bone marrow MSCs (hBM-MSCs), our team carried out lentiviral transduction of these cells for the evaluation of bioluminescent images (BLIs) in vitro and in vivo. In addition, MNPs that were previously characterized (regarding hydrodynamic size, zeta potential, and optical properties), and were used to label these cells, analyze cell viability via the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay and BLI analysis, and quantify the internalization process and iron load in different concentrations of MNPs via magnetic resonance imaging (MRI), near-infrared fluorescence (NIRF), and inductively coupled plasma-mass spectrometry (ICP-MS). In in vivo analyses, the same labeled cells were implanted in a sham group and a stroke group at different times and under different MNP concentrations (after 4 h or 6 d of cell implantation) to evaluate the sensitivity of triple-modal images.

Results: hBM-MSC collection and isolation after immunophenotypic characterization were demonstrated to be adequate in hBM samples. After transduction of these cells with luciferase (hBM-MSC_Luc), we detected a maximum BLI intensity of 2.0 x 10⁸ photons/s in samples of 10⁶ hBM-MSCs. Analysis of the physicochemical characteristics of the MNPs showed an average hydrodynamic diameter of 38.2 ± 0.5 nm, zeta potential of 29.2 ± 1.9 mV and adequate colloidal stability without agglomeration over 18 h. The signal of iron load internalization in hBM-MSC_Luc showed a close relationship with the corresponding MNP-labeling concentrations based on MRI, ICP-MS and NIRF. Under the highest MNP concentration, cellular viability showed a reduction of less than 10% compared to the control. Correlation analysis of the MNP load internalized into hBM-MSC_Luc determined via the MRI, ICP-MS and NIRF techniques showed the same correlation coefficient of 0.99. Evaluation of the BLI, NIRF, and MRI signals in vivo and ex vivo after labeled hBM-MSC_Luc were implanted into animals showed differences between different MNP concentrations and signals associated with different techniques (MRI and NIRF; 5 and 20 µg Fe/mL; P < 0.05) in the sham groups at 4 h as well as a time effect (4 h and 6 d; P < 0.001) and differences between the sham and stroke groups in all images signals (P < 0.001).

Conclusion: This study highlighted the importance of quantifying MNPs internalized into cells and the efficacy of signal detection under the triple-image modality in a stroke model.

Keywords: Bioluminescence; Human bone marrow mesenchymal stem cells; Magnetic resonance image; Multimodal nanoparticles; Near-infrared fluorescence image; Stroke.